Arsenic and trichloroethene (TCE) are among the most prevalent groundwater contaminants in the United States. Cocontamination of these two compounds has been detected at 63% of current TCE-contaminated National Priorities List sites. When in situ TCE reductive dechlorination is stimulated by the addition of fermentable substrates to generate a reducing environment, the presence of arsenic can be problematic because of the potential for increased mobilization and toxicity caused by the reduction of arsenate [As(V)] to arsenite [As(III)]. This study assesses the effects of arsenic exposure on the TCE-dechlorinating activities of Dehalococcoides mccartyi strain 195. Our results indicate that 9.1 μM As(III) caused a 50% decrease in D. mccartyi cell growth. While As(V) concentrations up to 200 μM did not initially impact TCE dechlorination, inhibition was observed in cultures amended with 200 μM As(V) and 100 μM As(V) in 12 and 17 days, respectively, corresponding with the accumulation of As(III). Transcriptomic and metabolomic analyses were performed to evaluate cellular responses to both As(V) and As(III) stress. Amendment of amino acids enhanced arsenic tolerance of D. mccartyi. Results from this study improve our understanding of potential inhibitions of D. mccartyi metabolism caused by arsenic and can inform the design of bioremediation strategies at co-contaminated sites.
Recovery and cultivation of diverse environmentally-relevant microorganisms from the terrestrial subsurface remain a challenge despite recent advances in modern molecular technology. Here, we applied complex carbon (C) sources, i.e., sediment dissolved organic matter (DOM) and bacterial cell lysate, to enrich groundwater microbial communities for 30 days. As comparisons, we also included enrichments amended with simple C sources including glucose, acetate, benzoate, oleic acid, cellulose, and mixed vitamins. Our results demonstrate that complex C is far more effective in enriching diverse and distinct microorganisms from groundwater than simple C. Simple C enrichments yield significantly lower biodiversity, and are dominated by few phyla (e.g., Proteobacteria and Bacteroidetes), while microcosms enriched with complex C demonstrate significantly higher biodiversity including phyla that are poorly represented in published culture collections (e.g., Verrucomicrobia, Planctomycetes, and Armatimonadetes). Subsequent isolation from complex C enrichments yielded 228 bacterial isolates representing five phyla, 17 orders, and 56 distinct species, including candidate novel, rarely cultivated, and undescribed organisms. Results from this study will substantially advance cultivation and isolation strategies for recovering diverse and novel subsurface microorganisms. Obtaining axenic representatives of “once-unculturable” microorganisms will enhance our understanding of microbial physiology and function in different biogeochemical niches of terrestrial subsurface ecosystems.
22The recovery and cultivation of diverse field-related microorganisms from the terrestrial 23 subsurface environment remains a challenge despite recent advances in modern 24 molecular technology. Here we applied natural organic carbon (C), i.e., sediment-derived 25 natural organic matter (NOM) and bacterial cell lysate, to groundwater microbial 26 communities for a 30-day enrichment incubation, followed by conventional direct-plating 27 for isolation. The groundwater was collected from a background well at the Oak Ridge 28Reservation Field Research Center, Tennessee. As a comparison, we also included 29 enrichments amended with simple organic C sources, including glucose, acetate, 30 benzoate, oleic acid, cellulose, and mixed vitamins. Our results demonstrate that complex 31 natural organic C sources are more effective in enriching diverse bacterial species from 32 groundwater than simple organic C sources. Microcosms amended with simple organic C 33 (glucose, acetate, benzoate, or oleic acid) show significantly lower biodiversity than 34 unamended control and are dominated by only few phyla such as Proteobacteria and 35Bacteroidetes. In contrast, microcosms amended with complex natural organic C 36 (sediment NOM or bacterial cell lysate) display significantly higher biodiversity, and 37 enrich distinct species from the phyla that are poorly represented in published culture 38 collections (e.g., Verrucomicrobia, Planctomycetes, and Armatimonadetes). Our 39 subsequent isolation efforts from natural organic C-amended enrichments led to 222 40 purified bacterial isolates representing 5 phyla, 16 orders, and 54 distinct species 41 including candidate novel, rarely cultivated, and undescribed organisms. 42 43 Importance 44Innovative strategies for recovering bacterial strains representing the true diversity of 45 microbial communities in the terrestrial subsurface would significantly advance the 46 understanding of ecologically critical taxa residing in these ecosystems. In this study, we 47 demonstrate that complex natural organic C that mimic the naturally available resources 48 for microbes encourages the growth of diverse bacteria much more robustly than 49 traditional simplistic organic C sources. Results from this study will substantially 50 advance and improve the design of strategies to effectively cultivate and isolate diverse 51 and novel subsurface microorganisms in the laboratory. Obtaining axenic cultures of the 52 'once-unculturable' microorganisms will greatly enhance our understanding of microbial 53 physiology, function, and roles in different biogeochemical niches in terrestrial 54 subsurface ecosystems. 55 56 57Compared to animal and plant hosts, other non-human environments on Earth 58 such as marine sediment, seawater, soil, and the terrestrial subsurface host prodigious and 59 undiscovered microbial populations, as most of them have never been cultured and 60 characterized in the laboratory 1 . In the terrestrial subsurface, it is estimated that there are 61 2,500 × 10 26 microbial cells, of whi...
Niche environmental conditions influence both the structure and function of microbial communities and the cellular function of individual strains. The terrestrial subsurface is a dynamic and diverse environment that exhibits specific biogeochemical conditions associated with depth, resulting in distinct environmental niches. Here, we present the characterization of seven distinct strains belonging to the genus Arthrobacter isolated from varying depths of a single sediment core and associated groundwater from an adjacent well. We characterized genotype and phenotype of each isolate to connect specific cellular functions and metabolisms to ecotype. Arthrobacter isolates from each ecotype demonstrated functional and genomic capacities specific to their biogeochemical conditions of origin, including laboratory-demonstrated characterization of salinity tolerance and optimal pH, and genes for utilization of carbohydrates and other carbon substrates. Analysis of the Arthrobacter pangenome revealed that it is notably open with a volatile accessory genome compared to previous pangenome studies on other genera, suggesting a high potential for adaptability to environmental niches.
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